In conditions of insulin resistance such as obesity and type 2 diabetes, women display a substantially increased risk for cardiovascular disease (CVD). As the lifetime risk for women in the United States to become diabetic reaches 40%, the impact of CVD in women continues to become a major health problem. In industrialized countries, the increased consumption of a high fructose and high fat diet (named Western Diet in this proposal) is a major driver for development of obesity, insulin resistance and type 2 diabetes. Insulin resistance in the vasculature results in decreased bioavailable nitric oxide (NO), impaired endothelial dependent dilation and is associated with increased vascular stiffness. Reduced NO results in increased activity of transglutaminase 2, an enzyme that increases collagen cross-linking and vascular stiffness. Importantly, vascular stiffness is a strong predictor of CVD and this abnormality is especially prevalent in obese, insulin resistant and diabetic women. Women with insulin resistance lose vascular protection normally afforded by estrogen signaling via estrogen receptor alpha (ER?), and our preliminary data suggest that in insulin resistant conditions, ER? signaling paradoxically contributes to the promotion of vascular stiffness in females. Furthermore, our results in a female rodent model of insulin resistance induced by Western Diet also demonstrate that mineralocorticoid receptor (MR) blockade improves vascular insulin resistance and stiffness. The role of the ER?-MR interaction in the genesis of vascular disease, particularly vascular stiffness, in insulin resistant females hs not been explored. Accordingly, the central hypothesis of this proposal is that in females, Western Diet-induced vascular insulin resistance and stiffness result from an interaction of endothelial cell MR and ER? signaling leading to reduce NO availability and increased TG2 activation. My proposal will use novel rodent models of endothelial specific MR and ER? knockout fed a Western Diet, as well as innovative techniques to access vascular stiffness in vivo and ex vivo. In Aim 1, I will determine mechanisms underlying the ER? and MR interaction as it relates to impaired insulin metabolic signaling and NO bioavailability in the vasculature of females fed a Western Diet. In Aim 2, I will determine the role of ER? and MR activation in the genesis of Western Diet induced vascular stiffness. In Aim 3, I will examine sexual dimorphic vascular stiffness effects of endothelial MR activation in the presence of intact ER? and WD-induced insulin resistance. I anticipate that results from this project will yield unique insights nto the mechanisms of vascular disease in obese and type 2 diabetic women, with the ultimate goal of translating these findings into therapeutic strategies to reduce CVD in vulnerable women.